Method for combatting bacteria using certain 2-(2&#39;-hydroxyphenyl)-oxazoles

ABSTRACT

A METHOD FOR COMBATTING HARMFUL MICROORGANISMS, SUCH AS FUNGI AND BACTERIA, BY APPLYING A 2-(2&#39;&#39;-HYDROXYPHENYL)OXAZOLE.

United States Patent Office Patented Dec. 19, 1972 US. Cl. 424-272Claims ABSTRACT OF THE DESCLOSURE A method for combatting harmfulmicroorganisms, such as fungi and bacteria, by applying a2-(2'-hydroxyphenyl)- oxazole.

CROSS-REFERENCE TO RELATED APPLICATION This is a continuation-in-part ofour application Ser. No. 686,298, filed Nov. 28, 1967, now abandoned.

BACKGROUND OF THE INVENTION The present invention provides a method forcombating harmful microorganisms, which comprises applying to a desiredsite an effective amount of a 2-(2'-hydroxyphenyl)- oxazole of theformula N i R CX or an alkali metal salt thereof, where R is an aromaticring system containing at most 2 six-membered rings, which is fused onas indicated by the valency lines, and X represents a benzene residuesubstituted by a hydroxyl group in ortho-position relatively to the bondwith the oxazole ring.

Of special value are 2-(2'-hydroxypheny1)-oxazoles of the formula inwhich R has the above meaning, Me is a hydrogen or alkali metal atom,and at least one of the symbols X and X represents a halogen atom, anamino group or an alkyl group containing up to 4 carbon atoms and theother is a hydrogen or halogen atom or an alkyl group with up to 4carbon atoms. The present invention is specially concerned with a methodfor combating harmful microin which Me has the above meaning; at leastone of the symbols X and X stands for a halogen atom or an alkyl groupwith up to 4 carbon atoms and the other for a hydrogen or halogen atomor for an alkyl group with up to 4 carbon atoms, and one to four of thesymbols R to R represent hydrogen or halogen atoms and two of thesesymbols, not standing for hydrogen or halogen atoms, represent phenyl,alkyl, alkoxy, phenylalkyl, cyclohexyl, nitro, primary amino,alkylamino, acylamino, trihalomethyl, aminosulphonyl,methylaminosulphony groups or sulphonic acid groups (possibly in theform of their alkali metal salts); or two vicinal symbols R to Rtogether form the complement to a fused-on carbocycle, which ispreferably unsubstituted, and any alkyl carbon chains in thesubstituents contain at most 12 carbon atoms. Within the indicatedmeanings -R to R may be chosen independently of one another, that is tosay for instance that one and the same molecule may contain 1 to 4halogen atoms side by side with 0 to 3 hydrogen atoms and 0 to 2,preferably 0 to 1, of the other substituents of the kinds indicatedabove.

Preferred compounds are the 2-(2-hydroxyphenyl)- benzoxazoles of theformula in which one or both of the symbols R and R are hydrogen,chlorine or bromine atoms, methyl, methoxy or nitro groups and, when oneof these symbols does not satisfy this definition, it stands for aphenyl, cyclohexyl group or a phenylalkyl group containing up to 9carbon atoms; R represents a hydrogen or halogen atom; X a hydrogen,chlorine or bromine atom or a methyl group and X a chlorine or bromineatom.

In this group 2-(2-hydroxyphenyl)-benzoxazoles of the formula N HO X n Il o I O X n where R is a hydrogen, chlorine or bromine atom, an alkylgroup with 1 to 4 carbon atoms, a phenyl, cyclohexyl or methoxy group; Rrepresents a hydrogen, chlorine or bromine atom or a methyl group; R ahydrogen, chlorine or bromine atom, and X and X a chlorine or bromineatom eachare particularly valuable as active ingredients ofantimicrobial preparations.

More especially, 2-(2-hydroxyphenyl)-benzoxazoles of theformula N HO X,l Q

on X:

where R is a hydrogen, chlorine or bromine atom, a methyl, phenyl orcyclohexyl group, and where R R X and X have the above meaningsarepreferred because of their good antimicrobial activity. Extremelysatisfactory results in combating microorganisms have been obtained with2-(2'-hydroxyphenyl)benzoxazoles of the formula N HO Xa 53' where R andR each is a hydrogen, chlorine or bror mine atom or a methyl group, andR X and X have the above meanings. Specifically, the following compoundsmay be mentioned, for example:

/N\ HO $1 which display an excellent activity against microorganisms, ep c a y g i st bacteria.

4 The 2-(2-hydroxyphenyl)-oxazoles of the Formula 1 can be prepared byknown methods by cyclizing an orthohydroxyamino compound of the formulawhere R has the above meaningwith a functional derivative of a2-hydroxybenzene-l-carboxylic acid (HOOC-X) and, if desired, introducingfurther substituents, for example halogen atoms or sulphonic acidgroups, into the resulting oxazole compound and/or converting it into analkali metal salt thereof.

An especially unexpected feautre of the compound of the Formula 1 istheir broad antibacterial activity spectrum which, in the case of someof these oxazoles, extends to both Gram-positive and Gram-negativebacteria. In this connection, as far as their application is concerned,the absence of odours and colours in the compound of the Formulas l to11 is of special value.

The present invention includes also the use of the 2-(2'-hydroxyphenyl)-oxazoles in pest control quite generally. Theantimicrobial compounds may be used on a very broad basis, especiallyfor protecting organic substrates from infestation by destructive andpathogenic (including phytopathogenic) microorganisms. Accordingly, theaforementioned compounds may be used as preservatives or disinfectantsfor textile materials and other industrial products of many kinds, inplant protection, agriculture, veterinary medicine and cosmetics.

From among non-textile industrial products that can be protected withthe aid of the oxazoles the following random examples may be given:Textile assistants and improving agents, glues, binders, paints, dyeingand printing pastes and similar preparations based on organic orinorganic dyestuffs or pigments, including those which contain a shareof casein or other organic compounds. Likewise, wall and ceiling paints,for example those containing a proteinic dyestutf binder, may beprotected from infestation by pests by the addition of the newcompounds. Timber may likewise be protected.

Furthermore, the oxazoles may be used for producing preserving anddisinfecting finishes on fibres and textile materials, including bothnatural and synthetic fibres on which they produce a durable protectionagainst harmful (including pathogenic) microorganisms, for example fungiand bacteria. The compounds may be added before, during or after atreatment of such textile materials with other substances, for exampledyeing or printing pastes, dressing agents etc.

Textile materials treated in this manner are also protected against bodyodour (perspiration) caused by microorganisms.

The oxazoles may also be used as preservatives in the cellulose andpaper industry, inter alia for preventing the known slime formation inpaper-making machines due to microorganisms.

Furthermore, when the oxazoles are combined with surface-active,especially detergent, substances, there are ob tained detergents andcleansing agents having a excellent antibacterial or antimycotic effect.The compounds of the Formulae 1 to 11 can be incorporated, for example,with soaps, or combined with soap-free detergents or surfaceactivesubstances or they may be combined with mixtures of soaps and soap-freedetergents, and their antimicrobial activity is fully maintained in suchcombinations.

Cleansing preparations containing compounds of the formulae shown abovemay also be used in industry and the home, in the food industry, forexample in dairies, breweries or abattoirs. They may also be used asingred- 'ients of preparations used for cleansing or disinfectinghospitals or surgeries.

The effect of the oxazoles can also be utilized in pro vinding plasticmaterials with a preserving and disinfecting finish. When plasticizersare used it is advantageous to dissolve or disperse the antimicrobialcompound in the plasticizer and then to incorporate it with the plasticmaterial. It is advantageous to ensure that the additive is as evenly aspossible distributed in the plastic material. Plastic materials havingantimicrobial properties can be used for making utilitarian articles ofall kinds in which an activity against a wide variety of germs, forexample bacteria and fungi, is desirable, for example in doormats,bathroom curtains, seats, treads in swimming baths, wall coverings orthe like. By incorporation with waxes and polishing compositions, floorand furniture polishes are obtained that have a disinfecting activity.

The antimicrobially active substances can be applied to the textilematerials to be protected in a variety of ways, for example byimpregnating or spraying with solutions or suspensions containing theabove-mentioned compounds as active ingredients. Depending on theindividual use, the content of active substance may vary from 1 to 30 g.per litre of treatment liquor.

In most cases textile materials of synthetic or natural origin will beadequately protected from infestation by fungi and bacteria byapplication of a preparation containing 0.1% to 3% of active substance.The active substances mentioned above may be used in combination withother textile assistants such as dressing agents, anticrease finishingagents or the like.

The form in which the active substances of this invention are appliedmay be similar to the conventional formulations of pesticidalpreparations; for example, prepara tions containing the said activesubstances may, if desired, further contain additives such as vehicles,solvents, diluents, dispersants, wetting agents, adhesives or the likeand also other pesticides.

Unless otherwise indicated, parts in the following manufacturinginstructions and examples are by weight.

MANUFACTURING INSTRUCTIONS (A) A mixture of 20.7 parts of3,5-dichlorosalicylic acid, 12.3 parts of 2 amino-4-methylphenol and 100parts of polyphosphorio acid is stirred and heated for 3 hours at195-200 C. under nitrogen. The reaction mixture is then poured in a thinjet over 2000 parts of ice water and the resulting suspension is stirredfor 1 /2 hours. The compound A of the formula is filtered off, washedneutral with water and dried. Yield: 28 parts. Melting point: 158160 C.After purification by recrystallization from chloroform+petroleum etherthe compound melts at l60l 6l C.

(B) A mixture of 20.7 parts of 3,5-dichlorosalicylic acid, 14.4 parts of2-amino-4-chlorophenol and 0.4 part of aluminium chloride in 140 partsby volume of o-dichlorobenzene is mixed at 130 C. within 30 minutes witha solution of 12 parts of phosphorus trichlon'de in parts by volume ofo-dichlorobenzene. The reaction mixture is then refluxed for 2 hours andpoured over ice water. The aqueous layer is separated and the organicphase repeatedly washed with water. The o-dichlorobenzene is expelledwith steam, to leave the compound B of in a yield of 22 parts, meltingat 199201 C. After recrystallization from dioxane-l-methanol thecompound melts at 204-205 C.

(C) 32.0 parts of bromine in 50 parts by volume of glacial acetic acidare added at 50-60 C. within minutes to a solution of 21.1 parts of2-(2'-hydroxyphenyl)- benzoxazole in 150 parts by volume of glacialacetic acid. The resulting suspension is heated for 2 hours longer at 60C., then cooled and at 25 C. 200 parts of water are added. The compoundC of the formula 5) N HO 1'31 -Q l is filtered off, rinsed with waterand dried. Yield: 34 parts. After recrystallization fromdioxane-i-acetonitrile or from dimethylformamide the compound melts at203-204 C.

(D) 56.0 parts of the compound of the formula 6) /N OH Cl (=Compound Fin the Table) are stirred within 40 minutes into 130 parts by volume ofchlorosulphonic acid. The temperature of the mixture rises gradually to4550 C., and a clear solution is obtained which is heated for 2 hours at105-110 C. and then poured in a thin jet over ice water. Theprecipitated compound of the formula 7) OH Cl hours and then cooled to25 C., whereupon the compound of the formula NELHO S 1130/ 0 or settlesout in light-yellow tflakes melting at 226-228 C. Yield: 17 parts.

A mixture of 4.05 parts of the dimethylamine salt and 20 parts by volumeof 2 N sodium hydroxide solution is heated for 15 minutes on a waterbath, during which dimethylamine is given off. The solution is cooled to25 C. and acidified with 2 N hydrochloric acid. The precipitate isfiltered off and once recrystallized from aqueous alcohol, to yieldabout 2.6 parts of the compound D of the formula melting above 400 C.

(E) A solution of 6.29 parts of the compound of the Formula (13) in 100parts by volume of dimethylformamide is mixed at C. with a solution of0.8 part of sodium hydroxide in 3.0 parts by volume of water. The

heating bath is removed and 100 parts by volume of alcobegins to settleout slowly. The reaction mixture is cooled hol are added at 45 C. Thecompound E of the formula to C. and the product filtered off and dried;it melts above 400 C. Yield: 5.4 parts.

B C1 As described in manufacturing instructions A to E 5 above, the2-(2-hydroxyphenyl)benzoxazoles F to Z, AA

0 to BB of the following table can be prepared; they corre- 3 spond tothe .above Formula 3 and the symbols R to R 0 1 X and X have themeanings shown in the table.

Analysis Calculated Found Meltin Compound R1 R R3 R4 X1 X2 point, C C HN O H N H H C1 Cl 160-161 51.17 3.08 4. 76 57. 02 3.17 4. 75 H H C1 Cl204-205 49. 64 1. 92 4. 49. 84 1. 97 4. 44 H H Br Br 203-204 42. 31 1.91 3. 80 41. 53 1. 95 4. 05 S OaNa. H C1 C1 700 39.91 1.80 3. 5840.02 1. 93 3. 70 H H C1 C1 400 46. 40 1. 4. 16 46. 28 1. 43 4. 11 H HC1 C1 185. 5-186. 5 74 2. 52 5.00 55. 96 2. 31 4. 93 C1 H C1 Cl 218-21949 64 1. 92 4. 45 49. 74 1. 97 4. 38 H H C1 158. 5-159. 5 55 2. 52 5. 0055. 49 2. 59 5.04 CI H H 01 183-184 55 74 2. 52 5.00 55. 57 2. 53 4. 98H C1 C1 Cl 201-202 44 74 1. 44 4. 01 44. 78 1. 52 4. 27 H 01 H 01201-202 49. 64 1. 92 4. 45 49. 76 2.10 4. 42 H H H Br 174-176 48.11 21.7 4. 32 47. 97 2. 25 4.05 H H Br Br 217-219 38. 70 1. 49 3. 47 38. 67 1.65 3. 37 Cl H Br Br 224. 5-226. 5 38. 7 1. 49 3. 47 38. 71 1. 56 3. 42CH H C1 Cl 170-180 57.17 3.08 4. 76 57.35 3.16 4. 59 H H H Cl 154-15564.75 3.88 5. 39 64. 79 3. 83 5. 37 H H H Br 163-165 55. 29 3. 31 4. 6155. 08 3. 53 4. 84 CH; H C1 Cl 232-233 58. 46 3. 4. 55 58. 25 3. 66 4. HH 01 Cl 149-151 60. 73 4. 50 4.17 61.02 4. 52 4. 26 H H Br Br 157-158. 548.03 3. 56 3. 29 47. 77 3. 54 3. 42

H C1 Cl 181-182 64.07 3.11 3. 3. 3.

H C1 Cl 151-152 63.00 4.73 3.87 63.09 4.90 3.83

: mmj I X H $113 11 C1 01 165166 66.31 4.30 3.52 66.05 4. 17 3.71

Y 11 H N Oz 11 C1 Cl 293-294 48. 03 1. 86 8. 62 18. 1O 2 O0 8. 60 Z HCH3 0 H H C1 C1 169-171 54. 21 2. 93 4. 52 54. 33 3 O6 4. 42

AA H H CHaNH 1'1 01 01 254-255 45. O6 2. 7O 7. 51 45. 35 2 7. 38

CH C1 C1 C1 278-279 78 2. 41. 3. 71 48. O6 2. 44 3. 58 H H CH; 131-13274. 65 4. 92 6. 22 74. 53 5. 11 6. 19 H C1 B! B! 264-265 44. 74 1. 44 4.01 45. O4 1. 69 4. 04 H Cl Br B1 291-292 33. O5 0. 2. 97 33. 25 O. 83 3.02 11 H CH; 165. 6-166 5 64. 75 3. 88 5. 39 64. 54 4. 02 5.46 H H CH:128-129 5 64. 75 3. 88 5. 39 64. 85 3. 87 5. 31 H H B1 B1 206-207 34.86 1. 35 3. 13 34. 89 1. 42 3. 04 H H CH3 B1 163-164 56. 62 3. 80 4. 4056. 76 4. 03 4. 42 H H CH] BI 183-134 49. 66 2. 68 4. 14 49. 73 2. 77 4.11 H H Bl CH3 218-219 49. 66 2. 68 4. 14 49. 52 2. 62 4. 19 H H Bl CH;168. 5-169 5 56. 62 3. 80 4. 40 56. 61 4. 0O 4. 16 H H 01 CH: 155. 5-1565 65. 82 4. 42 5. 12 65. 67 4. 54 4. 89 H H Cl CH1 195-19 57. 17 3. 084. 76 57. 34 3. 17 4. 62 H H CH; 01 174-175 57. 17. 3. 98 4. 76 57. 072. 86 4. 98 H H CH: 01 160-161 65. 82 4. 42 5. 12 65. 84 4. 48 4. 99 H HB1 C1 160. 5-161 5 49. 66 2. 68 4. 14 49. 78 2. 84 4. 07 H H B1 C1 213.5-215 43. 49 1. 68 3. 90 43. 73 2. 05 3. 92

(RH! H H 01 Cl 156-151 64. 29 5. 91 3. 57 64. 3'1 6. 99 3. 36 z):C-CHs-I AT H (3H3 H H B1 B1 160. 5-161 52. 41 4. 82 2. 91 52, 2O 4. 88 2 79a)a-CCHn OF; H H C1 Cl 169-171 48. 1. 74 4. 02 29 16. 9 4. 23

CF H H 131 B1 192-193 5 38. 48 1. 38 3. 21 38. 56 1. 41 3. 38 OF; H H B1140. 5-141 5 46. 1. 97 3. 91 47. 10 2. 11 3. 77 CF; H H Cl -146 53. 612. 25 4. 47 53. 76 2. 34 4. 28 $0311 H C1 Cl 327-329 43. 35 1. 96 3. B943. 12 2. 9O 4. 06 N0 H H C1 C1 223; 5-224 5 43. 03 1. 86 8. 62 47. 812. 04 8 60 NH; H H 01 Cl 265-266 52. 91 2. 73 9. 49 53. 12 2. 93 9. 72NH: H C1 Cl 273-274 52. 91 2. 73 9. 49 53. U9 2. 73 9. 49

The following compounds are accessible in an analogous manner:

Analysis Calculated Found Melting Compound Formula point, C. C H N C H NI /N\ H0 31 l o 01 BE 01 N\ H( 272-273 59.90 3.48 10.76 59.77 3.77 10.53

EXAMPLE 1 EXAMPLE 2 Determining the minimal inhibitory concentration(MIC) against bacteria in the dilution test The MIC (minimal inhibitoryconcentration) is determined by a method adapted from the standardprocedure, which furnishes an approximation to absolute minimalinhibitory values of an active substance.

A solution of 1% and 0.3% strength each of the above substances indimethylsulphoxide are poured into small tubes containing glucose brothand with these solutions dilution series are prepared in which eachmember is a tenfold dilution of the preceding one. By combining the twoseries, the following continuous dilution series is obtained: 1000, 300,100, 30, 10, 3 parts per million and so The solutions are inoculatedwith Staphylococcus aureus and then incubated for 48 hours at 37 C.(bacteriostatic test).

After the time mentioned, the minimal inhibitory values in parts permillion, shown in the following table, are found:

Compound: Minimal inhibitory concentration (MIC) l 111 parts per millionbacteriostatie test, Staphylococcus aureus.

Antibacterial soaps in cake form are manufactured by adding 1.2 g. of acompound of the Formula 1 to the following mixture:

120 g. of basic soap in flake form 0.12 g. of the disodium salt ofethylenediamine tetraacetic acid (dihydrate) 0.24 g. of titanium dioxide6 g. ethyleneglycol 10, 3, 1, and so on, parts per million of activesubstance.

The solutions are inoculated with the bacteria Staphylococcus aureus,Escherichia coli, and incubated for 24 hours at 37 C. After this time,0.05 ml. is withdrawn with a pipette from each solution and poured outover brain heart infusion slant-agar. 'Ihe agar tubes are then incubatedfor another 24 hours at 37 C., and then the minimal lethal concentrationin parts per million is determined.

Staphylococ- Escherichiiq 2 Compound cus aureus co EXAMPLE 3 Specimensof a Woolen fabric are impregnated with an 0.1 solution of a compound ofthe Formula I in dioxane and then padded to a liquid content of 100%.The dried fabric specimens contain 0.1% active substance, referred totheir own weight.

To test the effect against bacteria round blanks of mm. diameter of theimpregnated fabrics are placed on glucose agar plates, which havepreviously been inoculated with Staphylococcus aureus, and the platesare then incubated for 24 hours at 37 C.

The valuation extends, on one hand, to the inhibitory zones (IZ in mm.)found around the blanks and, on the other hand, to the growth (6%)detected underneath them by microscopic examination:

Staphylococcus aureus Compound IZ (mm.) percent What is claimed is:

1. A method for combatting bacteria which comprises applying to saidbacteria an anti-bacterially effective amount of a2-(2-hydroxyphenyl)-benzoaxole of the formula in which R represents amember selected from the group consisting of hydrogen, chlorine,bromine, alkyl with up to 4 carbon atoms, phenyl, cyclohexyl andmethoxy, R represents a member selected from the group consisting ofhydrogen, chlorine, bormine and methyl and R is hydrogen, cholrine orbromine and X and X represent chlorine or bromine each.

2. The method of claim 1, wherein the 2-(2'-hydroxyphenyl)-benzoxazolehas the formula.

l N HO X8 12 in which R represents a member selected from the groupconsisting of hydrogen, chlorine, bromine, methyl, phenyl andcyclohexyl.

3. The method of claim 1, wherein the 2'(2'-hydroxyphenyl) -benzoxazolehas the formula 4. The method of claim 3, wherein the2-(2'-hydroxyphenyl)-benzoxazole has the formula N H0 or 5. The methodof claim 3, wherein the 2-(2'-hydroxyphenyl)-benzoxazole has the formulaHO C1 References Cited FOREIGN PATENTS 957,438 5/ 1964 Great Britain.372,871 10/1963 Switzerland. 1,000,819 1/1967 Germany.

217,997 11/1961 Austria 260-307 ALBERT T. MEYERS, Primary Examiner L.SCHENKMAN, Assistant Examiner 5-6062/E/CONT mgg UNITED STATES PATENTOFFICE CERTIFICATE OF CORRECTION Patent No. 3 7 6, 3u Dated December 91972 Inventbfl) MAX SCHELLENBAUM ET AL It is certifiedthat error appearsin the above-identified patent and that said Letters Patent are herebycorrected as shown below:

Column 1, lines 5 and 6, delete "Corporation" and insert AG Signedandsealed this 2nd day of July 1974.

(SEAL) Attest: v

EDWARD MTFLETC HERJR. C. MARSHALL DANN Attesting Officer Commissioner ofPatents

